Hyperfibrinogenemia and sphingolipid-mediated cerebrovascular permeability and memory impairment during TBI

TBI期间高纤维蛋白原血症和鞘脂介导的脑血管通透性和记忆障碍

基本信息

批准号：
10855710
负责人：
DAVID LOMINADZE
金额：
$ 75万
依托单位：
UNIVERSITY OF SOUTH FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-08-15 至 2024-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10855710
关键词：
Address Alzheimer&apos s Disease Alzheimer&apos s disease related dementia Alzheimer&apos s disease risk Astrocytes Attenuated Avidity Binding Blood Blood Proteins Blood Vessels Brain Caveolae Complex Confocal Microscopy Cortical Contusions Crossbreeding Data Dementia Deposition Development Disease Electron Microscopy Electrons Endothelial Cells Endothelium Enzymes Exocytosis Extravasation Fibrinogen Fluorescence Fluorescence Resonance Energy Transfer Fumonisins Generations Glucosylceramides Goals Heart Diseases Immunohistochemistry Impaired cognition In Situ In Vitro Inflammation Injury Intercellular adhesion molecule 1 Knockout Mice MAPK3 gene Measures Mediating Memory Memory Loss Memory impairment Methods Microscopy Mitochondria Modeling Morbidity - disease rate Mus N-palmitoylsphingosine Pathogenicity Pathway interactions Peripheral Nervous System Permeability Phosphorylation Pilot Projects PrP Production Proteins Research Research Priority Resistance Role Short-Term Memory Signal Transduction Small Interfering RNA Sphingolipids Sphingomyelins Testing Tracer Transgenic Organisms Traumatic Brain Injury United States National Institutes of Health Vascular Cognitive Impairment Vascular Diseases Vascular Endothelial Cell blood-brain barrier permeabilization brain endothelial cell caveolin 1 cerebrovascular dihydroceramide desaturase in vivo inhibitor insight knockout gene loss of function mortality mouse model neuroinflammation new therapeutic target novel object recognition protein complex receptor serine palmitoyltransferase thermozymocidin transcytosis

项目摘要

Project Summary Increased blood content of fibrinogen (Fg), e.g. hyperfibrinogenemia (HFg) is a risk factor for Alzheimer's disease (AD). As it occurs during neuroinflammation, it is a risk factor for AD & related disorders (ADRD), including vascular cognitive impairment & dementia (VCID). Our goal is to characterize the role of a novel mechanistic pathway, namely Fg-sphingolipid-caveolae nexus during traumatic brain injury (TBI) as an example of ADRD, which is accompanied with HFg. Our data indicate that cortical contusion injury (CCI)-induced HFg enhances cerebrovascular permeability mainly via caveolar protein transcytosis leading to Fg deposition in extravascular space and resulting in greater formation of Fg and cellular prion protein complexes resulting in short-term memory (STM) reduction, typically occurring in AD. We showed that HFg increased Fg binding to endothelial intercellular adhesion molecule-1 (ICAM-1) enhancing formation of functional caveolae & exocytosis via activation of ERK-1/2 and caveolin-1 (Cav-1). We found that HFg upregulated de novo sphingolipid synthesis pathway & caveolar transcytosis using mitochondrial ATP. A selective inhibitor of sphingolipid synthesis pathway ameliorated the HFg- triggered caveolar protein transcytosis in mouse brain endothelial cells, & importantly, reduced cerebrovascular protein transcytosis after CCI. Based on these results, we hypothesize that at elevated levels, Fg, through binding to its endothelial receptor ICAM-1, activates sphingolipid production resulting in increased caveolar protein transcytosis in ECs that is supported by ATP generated in mitochondria. A corollary hypothesis is that an enhanced caveolar Fg transcytosis contributes to STM reduction similar to that during AD. Specific aims are: (1) Determine whether the increased interaction of Fg with endothelial ICAM-1 enhances formation of functional caveolae via production of Cer, GlcCer, and SPM sphingolipids and thereby increases caveolae-mediated protein transcytosis. (2) Determine whether increased interaction of Fg with ECs enhances mitochondrial activity using up energy for caveolar protein transcytosis. (3) Determine whether inhibition of de novo sphingolipid synthesis attenuates HFg- induced increased formation of functional caveolae decreasing caveolar protein transcytosis and thus, ameliorates the STM reduction during TBI. To test mechanisms of HFg-induced caveolar transcytosis via sphingolipid signaling we will use WT, HFg, and endothelium-specific serine palmitoyltransferase long chain-2 gene knockout (Sptlc2endo-/-) and HFg/Sptlc2endo-/- mice generated by Cre/lox method, with or without CCI. The dual-tracer probing method will be used to define changes in caveolar transcytosis. Loss-of-function strategy (specific siRNAs against ICAM-1 & Cav-1) and specific inhibitors of sphingolipid synthesis will be used. Immunohistochemistry and intravital, confocal, electron, total internal reflection fluorescence, and fluorescence resonance energy transfer microscopies will be used. Changes in STM will be assessed by a novel object recognition and Y-maze tests. These studies will provide insight into the role of sphingolipids in mechanisms of cerebrovascular permeability & may unveil new therapeutic targets for one of the leading causes of VCID.

项目摘要纤维蛋白原（FG）的血液含量增加，例如高纤维蛋白酶血症（HFG）是阿尔茨海默氏病的危险因素（广告）。由于在神经炎症期间发生，它是广告和相关疾病的危险因素（ADRD），包括血管认知障碍和痴呆（VCID）。我们的目标是表征新机械的作用途径，即创伤性脑损伤（TBI）期间FG-Sphingolipid-Caveolae Nexus，作为ADRD的一个例子伴随着HFG。我们的数据表明皮质挫伤损伤（CCI）诱导的HFG增强了脑血管渗透性主要通过洞穴蛋白转胞菌病，导致血管外FG沉积空间和导致FG和细胞prion蛋白复合物的形成更大，导致短期记忆（STM）还原，通常发生在AD中。我们表明，HFG增加了FG与内皮间细胞间的结合通过激活ERK-1/2的激活，粘附分子-1（ICAM-1）增强功能性小窝和外吞作用的形成和小窝1（CAV-1）。我们发现，HFG上调了新的鞘脂合成途径和Caveolar 使用线粒体ATP的转介症。鞘脂合成途径的选择性抑制剂改善了HFG- 在小鼠脑内皮细胞中触发了小肠蛋白转胞细胞增多症，重要的是，脑血管减少 CCI后蛋白质转胞病。基于这些结果，我们假设在升高的水平，Fg，通过结合至其内皮受体ICAM-1，激活鞘脂的产生，导致洞穴蛋白增加在线粒体中产生的ATP支持的EC中的转胞病。推论假设是增强 Caveolar FG转胞菌病有助于与AD相似的STM还原。具体目的是：（1）确定 FG与内皮ICAM-1的相互作用增加是否增强了功能性小窝的形成 CER，GLCCER和SPM鞘脂的产生，从而增加小窝介导的蛋白质转胞病。（2）确定FG与EC的相互作用是否增加会增强使用UP能量的线粒体活性洞穴蛋白转介作用。（3）确定对从头鞘脂合成的抑制是否会减弱HFG- 诱导的功能性小窝的形成增加，减少了小窝蛋白转胞菌病，因此可以改善 TBI期间的STM降低。测试HFG诱导的Caveolar transctosos通过鞘脂的机理信号我们将使用WT，HFG和内皮特异性丝氨酸棕榈酰转移酶长链2基因敲除（SPTLC2ENDO - / - ）和HFG/SPTLC2ENDO - / - 由CRE/LOX方法产生的，带有或不带CCI。双跟踪探测方法将用于定义洞穴跨介症的变化。功能丧失策略（针对的特定siRNA 将使用ICAM-1和CAV-1）以及鞘脂合成的特定抑制剂。免疫组织化学和插入术，共聚焦，电子，总内反射荧光和荧光共振能量转移显微镜将使用。 STM的变化将通过新颖的对象识别和Y迷宫测试来评估。这些研究会提供有关鞘脂在脑渗透性机制中的作用的见解，并可能揭示新的 VCID主要原因之一的治疗靶标。